In the past year, Zika virus has gained worldwide attention due to the rapidly spreading infection, its association with Guillain-Barr syndrome, its ability to be transmitted sexually and in utero to the fetus, and the increased number of infants with microcephaly and other severe brain defects. The genome of the current epidemic virus has 75 amino acid substitutions relative to the virus originally isolated from a rhesus monkey in Uganda in 1947 (MR766), which appears to have caused only sporadic and mild disease. Whether any of these substitutions has effects on viral replication fitness, pathogenesis, or spread between humans is unknown. Thus it is important that studies be done using the African isolate as well as more recent epidemic strains. The goal of this proposal to elucidate the underlying mechanisms governing the severe neural pathogenesis of the epidemic virus and to test our hypothesis that induction of autophagy will inhibit the virus infection. This proposal brings together the knowledge of two well-established and experienced investigators in related fields. By combining the expertise in the Spector lab in virology, specifically the molecular/cell biology and pathogenesis of human cytomegalovirus (currently the major viral cause of neural birth defects), and stem cell differentiation, and in the Yeo lab on human stem cell biology (including differentiation of human induced pluripotent stem cells [iPSCs] to cerebral cortex neurons), high-throughput analysis of gene expression and RNA processing (including single cell RNA seq), and cutting edge computational technologies, we have a unique opportunity to gain greater understanding of this major medical problem. We will determine the kinetics of viral infection and cell death when cells are infected at different stages of differentiation of iPSCs to cortical neurons. We will also leverage our strengths in genome-wide technologies to identify changes in expression of host genes (including coding and noncoding RNAs and microRNAs) during infection. Importantly, we will compare the differences between infection with the early African Zika virus strain MR766 and a more recent epidemic isolate. Also novel and potentially of great therapeutic significance is our proposed strategy of inducing autophagy to inhibit ZIKV infection and promote cell survival and differentiation. Specifically, we will test whether treatment of cells with the nontoxic disaccharide trehalose, which induces autophagy via a novel mTOR-independent pathway, will inhibit the infection, as we have found for human cytomegalovirus. We will also test 2 other autophagy inducers, a mTOR-independent small molecule SMER28 and tat-beclin 1. Accomplishment of the goals of this proposal will facilitate the development of new strategies designed to prevent and treat Zika virus infection.